中国仓鼠卵巢细胞
泰特
生存素
分子生物学
四环素
细胞培养
生物
细胞凋亡
化学
基因
基因表达
抑制因子
遗传学
抗生素
作者
Xinghui Zhao,Yanping Yu,Zhongchao Zhao,Jia Guo,Ling Fu,Yasuyuki Takata,Lihua Hou,Shaoqiong Yi,Wei Chen
摘要
An optimized method based on tetracycline-inducible gene expression system T-REx was developed to screen and evaluate Tet repressor (TetR)-expressing cell lines using enhanced green fluorescence protein (EGFP) as reporter gene. To verify the effectiveness of the method, two TetR-expressing Chinese hamster ovary (CHO) cell lines, CHO-TR B2 (stringent) and B5 (less stringent), in which the EGFP genes were variantly controlled by tetracycline, were used to construct cell lines expressing the anti-apoptosis gene survivin upon induction with tetracycline. The resulting stable clones were analyzed for survivin expression. The analysis showed that all four B5-derived clones exhibited leaky survivin expression in the absence of tetracycline, while the B2-derived clones did not. DNA laddering and annexin V/PI staining assays further indicated that although tetracycline-inducible expression of survivin conferred resistance to NH4Cl- and staurosporine-induced apoptosis in both the B2- and the B5-derived stable cell lines, the B2-derived cell lines showed more stringent regulation in the absence of tetracycline. This represents successful utilization of the present screening method.
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